The enteric nervous system (ENS) is large, complex, and capable of integrative neuronal activity. Unlike the CNS, however, the ENS is relatively exposed in the abdomen to injury. Enteric neurons, moreover, have to withstand constitutive stresses of deformation and are potentially threatened by the microbiome to which the gut plays host. The ENS must work together with innate and adaptive immune mechanisms to maintain the sterility of the bowel wall. Although maintenance, plasticity, and renewal would thus seem to be important, relatively little is known about how they operate in the ENS. Serotonin (5-HT) has been implicated in neurotransmission, paracrine initiation of peristaltic and secretory reflexes, and nociceptive signaling to the CNS. Recent observations suggest that 5-HT and the 5-HT4 receptor subtype also function in ENS neuroprotection/neurogenesis and plasticity. These new data are compatible with the ideas that stimulation of 5-HT4 receptors promotes the survival of adult enteric neurons and also their generation from neuronally competent stem cells. We have found that the severity of 3 different forms of experimental intestinal inflammation is increased in mice that lack the serotonin transporter (SERT). Because the effects of 5-HT are potentiated in these animals, the observations suggest that 5-HT promotes intestinal inflammation. We now propose to test the hypotheses that 5-HT plays 2 different, but synergistic roles in defense of the gut: (i) Mucosal 5-HT, synthesized by tryptophan hydroxylase1 (TpH1), promotes inflammation (a function that might help to protect the bowel from microbial invasion). (ii) Neuronal 5-HT, synthesized by TpH2, is neuroprotective, promotes plasticity, particularly of dopaminergic neurons, and stimulates neurogenesis. These functions putatively protect the ENS from inflammation. We will now test these hypotheses with gain-of-function (SERTnull) and loss-of-function models (mice in which 5-HT biosynthesis is selectively prevented in the mucosa [TpH1null], the ENS [TpH2null], or both [double knockout, TpH1null + TpH1null]). These animals will make it possible to evaluate the relative importance of mucosal or neuronal sources of 5-HT in enteric neuroprotection, neurogenesis, neuronal plasticity and inflammation. Specifically, we will determine whether endogenous 5-HT is essential for survival of enteric neurons, avoiding degeneration of neurites, and recruiting precursors to generate new neurons, whether these actions are 5-HT4-mediated (utilizing 5-HT4null mice), and whether they depend on neuronal or mucosal 5- HT. We will also analyze the responsible signal transduction pathways, investigate the roles played by neuronal and mucosal 5-HT in enteric dopaminergic neuronal maintenance, and promotion of inflammation. Finally, we will determine whether selective inhibition of TpH1 can help to control intestinal inflammation. ) PUBLIC HEALTH RELEVANCE: Narrative The enteric nervous system (ENS; second brain) is complex and able to control the behavior of the bowel. Although the ENS is thus critical for life, it is also vulnerable to injury. The ability of the ENS to maintain and repair itself and adapt to change is thus important; nevertheless, relatively little is known about how the ENS accomplishes these tasks. Serotonin is known to be an important signaling molecule in the gut, serving both as a hormone and as a neurotransmitter. We have recently uncovered evidence that serotonin acts, in addition, to promote the survival of adult enteric neurons and their generation from stem cells. Serotonin also appears to be necessary for the maintenance of the subset of enteric neurons that use dopamine as their transmitter and has the further ability to promote intestinal inflammation. We now propose to test the hypotheses that there are two functional serotonin pools that defend the gut. One is a large pool synthesized by tryptophan hydroxylase1 (TpH1) that promotes inflammation, in part, to counter microbial invasion from the gut lumen and the other is a small neuronal serotonin pool, synthesized by TpH2, which protects neurons from inflammation and other causes of injury. We will now test these hypotheses with gain- of-function and loss-of-function models in which critical molecules have genetically been deleted in mice. We will also analyze the cellular receptors and signals through which the associated actions of serotonin are mediated. This work is relevant to the pathogenesis and treatment of inflammatory bowel disease, irritable bowel syndrome, and also to the disabling intestinal manifestations of Parkinson's disease. )